The overall objective of this proposal is to study in mammalian systems the role of DNA mismatch repair in genetic recombination and related DNA metabolic processes. To accomplish this goal we will characterize the structure and function of a mammalian gene (mmr1) that, based on DNA sequence, is highly homologous to a DNA mismatch repair gene conserved in bacteria and yeast. Subsequent to the structural analyses we will use, as a primary strategy, gene targeting protocols to perform gene disruption and to subsequently study the interplay between DNA mismatch repair, recombination and other aspects of DNA metabolism, e.g. mutagenesis, both in cultured cells and in laboratory mice. The specific aims are: 1) To characterize the structure and expression of the mammalian mmr1 gene. The main emphasis will be to gather sufficient genomic and cDNA derived information for rational gene disruption at this locus. 2) To disrupt the mmr1 gene in a near-euploid mouse cell line and study certain aspects of homologous chromosomal recombination, repair and mutagenesis in culture. 3) To disrupt the mmr1 gene in mouse embryonic stem (ES) cells and study the effects in laboratory mice. The proposed studies represent a logical extension of our previous studies of homologous recombination as it occurs within mammalian chromosomes. The studies are designed to provide information on mammalian DNA mismatch repair and the role it plays in homologous recombination and other DNA metabolic processes such as mutagenesis. In addition, this project should provide a number of useful tools, ranging from reagents for the analysis of mismatch repair at the biochemical level, to new and useful mutant strains of mice.